Summary/abstract Polycomb repressive complex 2 (PRC2) is an essential histone methyltransferase required for epigenetic silencing in development. Upregulation of PRC2 or its methyltransferase activity is frequently observed in cancer and can be critical for cancer cell proliferation and progression. Despite many efforts in reducing the expression level of PRC2 and inhibiting its catalytic active site in treatment of diseases, PRC2-nucleic acid interactions have been underestimated as important processes to regulate recruitment and catalytic activities of PRC2. It has been recently demonstrated by our group and others that RNA binding inhibits catalytic activities of PRC2, indicating that mutations of RNA binding residues of PRC2 subunits could be an alternative way to upregulate PRC2 activity and indeed such mutations have been observed in cancer (R34L, K39E, K491R and R494S in EZH2, the catalytic subunit of PRC2). Moreover, a few long noncoding (lnc)RNAs have emerged as key playmakers to recruit or evict PRC2 from chromatin loci to regulate specific gene expression in development and diseases, but understanding the physiological functions of RNA binding of PRC2 has been hampered previously by the lack of separation-of-function mutant, which has been finally identified in my recent publication (Long et al. eLife 2017). More recently, PRC2-DNA interaction has been shown to drive the interaction of PRC2 with nucleosome, and DNA-binding domains reside on multiple accessory subunits of PRC2 complex including AEBP2, JARID2, PHF1, MTF2 and PHF19. However, the genome-wide function of these DNA interactions has not been comprehensively investigated. To answer all these critical questions on PRC2 regulation, I propose a comprehensive in vitro and in vivo investigation of the molecular mechanisms and cellular functions of PRC2-nucleic acid interactions. Specifically, this proposal aims to: (1) understand how RNA interactions regulate cellular activities of PRC2; (2) understand how DNA regulates PRC2 through interaction with PRC2?s accessory proteins. During the K99 phase, under the mentorship of Dr. Tom Cech and Dr. John Rinn, comprehensive approaches using in vitro biochemical analysis and in vivo genome-wide characterization will be used to determine molecular mechanism and function of PRC2-nucleic acid interactions. With support from Dr. Leslie Leinwand, expertise in analyzing cardiomyocyte physiology and behavior will be acquired in order to identify important biological consequences of loss of nucleic acid interaction of PRC2 in cardiomyocytes, in which PRC2 activities have been shown to be critical. Additional K99 training in genomics and computational biology will be essential for the proposed research. All training above will facilitate continued investigations of functions of PRC2-nucleic acid interactions during the independent R00 phase. The results of this proposal will not only lead to identification of important PRC2-nucleic acid interactions and their molecular mechanisms, but also provide a paradigm for characterizing other epigenetic modifiers that bind nucleic acids especially noncoding RNAs.